The regulation of the permeability of the blood-brain barrier (BBB) is a dynamic process that includes the interaction between endothelial cells, the basement membrane and astrocytic end-feet processes. The onset of cerebral ischemia induces a complex set of changes in the BBB that leads to the passage of inflammatory cells from the intravascular space into the abluminal side of the endothelial cell-basement membrane- astrocyte interface associated with the development of cerebral edema, which is a leading cause of mortality in acute stroke patients (up to 80% in some series). Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) is a member of the tumor necrosis factor superfamily that acts on responsive cells via binding to a cell surface receptor known as fibroblast growth factor-inducible 14 (Fn14). We have previously demonstrated that in the brain TWEAK and Fn14 are expressed mainly in perivascular astrocytes and endothelial cells, and that the intracerebral injection of recombinant TWEAK induces NF-:B pathway activation in these cells and a dose-dependent increase in both matrix metalloproteinase-9 (MMP-9) activity and the permeability of the BBB. We have also found that following middle cerebral artery occlusion (MCAO) the expression of TWEAK and Fn14 increases in the ischemic area, and that inhibition of TWEAK activity following MCAO either by administration of a Fn14-Fc decoy receptor or by genetic deficiency of Fn14 results in inhibition of NF-:B activation and preservation of the structure and barrier function of the BBB. In consideration of these and other findings discussed below, in this application we hypothesize that during cerebral ischemia the interaction between TWEAK and Fn14 is a key molecular mechanism that regulates the permeability of the BBB. More specifically, we propose that early after the onset of the ischemic insult the binding of TWEAK to Fn14 in the endothelial cell- basement membrane-astrocyte interface results in the following sequence of events: i) NF-:B-mediated increased expression of the chemokine MCP-1 (CCL2) and its receptor CCR2 with the formation of a chemokine gradient that leads to the infiltration of inflammatory cells into the abluminal side of the BBB, ii) increase in inflammatory cell- derived MMP-9 activity, and iii) MMP-9-mediated increase in the permeability of the BBB with development of cerebral edema. In the last Aim of this proposal we will study whether inhibition of TWEAK-Fn14 binding with Fn14-Fc decoy delivered to the endothelial cell-basement membrane-astrocyte interface with a nanocarrier protects the barrier function of the BBB during ischemic conditions. If successful, these clinically relevant studies not only will shed light into the biology of this novel pathway but also may lead to the discovery of a new mechanism underlying the development of cerebral edema. Importantly, if our hypothesis is correct, our findings should indicate whether inhibition of TWEAK-Fn4 binding is a valid new target for the treatment of patients with acute ischemic stroke. PUBLIC HEALTH RELEVANCE: Early after the onset of ischemic stroke the interaction between the cytokine TWEAK and its receptor Fn14 induces brain swelling (edema), which is a very common cause of death among patients with ischemic stroke. The relevance of stroke among USA Veterans has been widely recognized. In fact, some studies have reported that the mortality due to ischemic stroke among USA Veterans is higher than in the general population, particularly in southern states of the country. In this application we study the mechanism whereby the interaction between TWEAK and Fn14 induces brain edema, and test the therapeutic efficacy of inhibitors of the TWEAK-Fn14 pathway for the treatment of acute ischemic stroke. Thus, if successful, the studies proposed in this application may lead to a decrease in the morbility and mortality among USA Veterans.